{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/22104"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/22104","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Covalent post-translational modification of proteins in Escherichia coli: Genetic and biochemical studies of biotin and lipoic acid biosynthesis and function","abstract":"I have identified four distinct classes of mutants involved in either lipoic acid biosynthesis or the covalent attachment of lipoic acid to proteins in E. coli. Two of these classes of mutants were isolated as Tn1000dKn insertion mutants and both classes map to min 14.5 on the E. coli chromosome. The other two classes of mutants were isolated as spontaneous selenolipoic acid resistant (slr) mutants. Selenolipoic acid was shown to be a potent inhibitor of wild type E. coli.","abstract_html":"I have identified four distinct classes of mutants involved in either lipoic acid biosynthesis or the covalent attachment of lipoic acid to proteins in E. coli. Two of these classes of mutants were isolated as Tn1000dKn insertion mutants and both classes map to min 14.5 on the E. coli chromosome. The other two classes of mutants were isolated as spontaneous selenolipoic acid resistant (slr) mutants. Selenolipoic acid was shown to be a potent inhibitor of wild type E. coli.","abstract_has_math":false,"creators":["Reed, Kelynne Elizabeth"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Microbiology","degree_department":null,"school":null,"contributors":["Cronan, John E."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2011,"date_issued":"2011-05-07T13:29:04Z","date_published":"2011-05-07T13:29:04Z","updated_at":"2026-07-22T22:25:19Z","subjects":["Biology, Microbiology"],"languages":["eng"],"rights":["Copyright 1992 Reed, Kelynne Elizabeth"],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9305663","(UMI)AAI9305663"],"render_values":[{"text":"AAI9305663","href":null,"code":true},{"text":"(UMI)AAI9305663","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2142/22104","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Cronan, John E."]},{"key":"dc:creator","label":"Author","values":["Reed, Kelynne Elizabeth"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2011-05-07T13:29:04Z","10000-01-01","1992"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Microbiology"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Dissertation"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Ph.D."]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["University of Illinois at Urbana-Champaign"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Biology, Microbiology"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 1992 Reed, Kelynne Elizabeth"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9305663","(UMI)AAI9305663","http://hdl.handle.net/2142/22104"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["I have identified four distinct classes of mutants involved in either lipoic acid biosynthesis or the covalent attachment of lipoic acid to proteins in E. coli. Two of these classes of mutants were isolated as Tn1000dKn insertion mutants and both classes map to min 14.5 on the E. coli chromosome. The other two classes of mutants were isolated as spontaneous selenolipoic acid resistant (slr) mutants. Selenolipoic acid was shown to be a potent inhibitor of wild type E. coli.","The Tn1000dKn mutations defined two genes, lipA and lipB, involved in the production of lipoylated proteins. Both the lipA and lipB genes were sequenced. The deduced amino acid sequence of the lipA gene showed some similarity to biotin synthase. It is thus likely that lipA encodes a sulfur insertion enzyme analogous to biotin synthase and that the two enzymes share a common sulfur donor. The slr-7 allele (maps to min 15.25) was shown to be a suppressor of lipA150::Tn1000dKn indicating that the slr-7 gene also plays a role in lipoic acid biosynthesis.","The amino acid sequence of the lipB gene did not share significant amino acid similarity to any protein in the Genebank Data Base. However, several experiments support the conclusion that the lipB gene is required for the covalent attachment of endogenously synthesized lipoic acid to protein. First, analysis of the lipoic acid content of a lipB null mutant grown under lipoate deficient conditions revealed that lipB mutants possessed functional, but attenuated lipoic acid biosynthetic activity. Secondly, the lipoic acid auxotrophic phenotype of a lipB null mutant could be suppressed by overexpression of the lut encoded lipoyl ligase (T. Morris, manuscript in preparation). The primary function of the lut encoded ligase is to covalently attach exogenous sources of lipoic acid to protein. Finally, I have shown that a lut lipB double mutant does not synthesize any detectable lipoyl-proteins.","I have shown that slr-1 is an allele of lut. Not only is the slr-1 mutant defective for lipoyl ligase activity, it is also 100 fold more resistant to selenolipoic acid than an isogenic wild type strain. Moreover, the slr-1 lipoyl ligase is able to distinguish between lipoic acid and selenolipoic acid indicating that slr-1 mutant is a specificity mutant.","Made available in DSpace on 2011-05-07T13:29:04Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9305663.pdf: 5255003 bytes, checksum: d71a562b391e50fc795d081b78f0a386 (MD5) Previous issue date: 1992","Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:55:21Z Item is restricted indefinitely.","Restriction data tranferred 2014-07-01T11:25:47-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission","ETDs are only available to UIUC Users without author permission","U of I Only"]},{"key":"dc:title","label":"Title","values":["Covalent post-translational modification of proteins in Escherichia coli: Genetic and biochemical studies of biotin and lipoic acid biosynthesis and function"]}]}],"canonical_facts":{"dc:contributor":["Cronan, John E."],"dc:creator":["Reed, Kelynne Elizabeth"],"dc:date":["2011-05-07T13:29:04Z","10000-01-01","1992"],"dc:description":["I have identified four distinct classes of mutants involved in either lipoic acid biosynthesis or the covalent attachment of lipoic acid to proteins in E. coli. Two of these classes of mutants were isolated as Tn1000dKn insertion mutants and both classes map to min 14.5 on the E. coli chromosome. The other two classes of mutants were isolated as spontaneous selenolipoic acid resistant (slr) mutants. Selenolipoic acid was shown to be a potent inhibitor of wild type E. coli.","The Tn1000dKn mutations defined two genes, lipA and lipB, involved in the production of lipoylated proteins. Both the lipA and lipB genes were sequenced. The deduced amino acid sequence of the lipA gene showed some similarity to biotin synthase. It is thus likely that lipA encodes a sulfur insertion enzyme analogous to biotin synthase and that the two enzymes share a common sulfur donor. The slr-7 allele (maps to min 15.25) was shown to be a suppressor of lipA150::Tn1000dKn indicating that the slr-7 gene also plays a role in lipoic acid biosynthesis.","The amino acid sequence of the lipB gene did not share significant amino acid similarity to any protein in the Genebank Data Base. However, several experiments support the conclusion that the lipB gene is required for the covalent attachment of endogenously synthesized lipoic acid to protein. First, analysis of the lipoic acid content of a lipB null mutant grown under lipoate deficient conditions revealed that lipB mutants possessed functional, but attenuated lipoic acid biosynthetic activity. Secondly, the lipoic acid auxotrophic phenotype of a lipB null mutant could be suppressed by overexpression of the lut encoded lipoyl ligase (T. Morris, manuscript in preparation). The primary function of the lut encoded ligase is to covalently attach exogenous sources of lipoic acid to protein. Finally, I have shown that a lut lipB double mutant does not synthesize any detectable lipoyl-proteins.","I have shown that slr-1 is an allele of lut. Not only is the slr-1 mutant defective for lipoyl ligase activity, it is also 100 fold more resistant to selenolipoic acid than an isogenic wild type strain. Moreover, the slr-1 lipoyl ligase is able to distinguish between lipoic acid and selenolipoic acid indicating that slr-1 mutant is a specificity mutant.","Made available in DSpace on 2011-05-07T13:29:04Z (GMT). No. of bitstreams: 2 license.txt: 4922 bytes, checksum: 910b249b4beec47e7ab768910c8f966f (MD5) 9305663.pdf: 5255003 bytes, checksum: d71a562b391e50fc795d081b78f0a386 (MD5) Previous issue date: 1992","Item marked as restricted to the 'UIUC Users [automated]' Group (id=2) by Howard Ding (hding2@illinois.edu) on 2011-05-07T14:55:21Z Item is restricted indefinitely.","Restriction data tranferred 2014-07-01T11:25:47-05:00 Original Data Group with Access UIUC Users [automated] Release Date: none Reason: ETDs are only available to UIUC Users without author permission","ETDs are only available to UIUC Users without author permission","U of I Only"],"dc:identifier":["AAI9305663","(UMI)AAI9305663","http://hdl.handle.net/2142/22104"],"dc:language":["eng"],"dc:rights":["Copyright 1992 Reed, Kelynne Elizabeth"],"dc:subject":["Biology, Microbiology"],"dc:title":["Covalent post-translational modification of proteins in Escherichia coli: Genetic and biochemical studies of biotin and lipoic acid biosynthesis and function"],"dc:type":["text"],"thesis:degree_discipline":["Microbiology"],"thesis:degree_level":["Dissertation"],"thesis:degree_name":["Ph.D."],"thesis:institution_name":["University of Illinois at Urbana-Champaign"]},"updated_at":"2026-07-22T22:25:19Z"}